PROJECT SUMMARY ? Core 3 The Correlative Studies and Immune Monitoring Core (Core 3) will provide all three projects comprising this Program Project Grant proposal comprehensive, state-of-the-art biorepository and immune/molecular profiling support by a team of highly accomplished research scientists who have utilized these specific platforms in conjunction with previous and/or current cancer immunotherapy trials. These three projects investigate the safety, efficacy and mechanisms of novel immunotherapeutic approaches to the treatment of patients with glioblastoma (GBM), a diagnosis with extremely poor prognosis. The objectives of this Core are to identify immunologic correlatives that predict clinical outcomes and provide mechanistic insights utilizing a comprehensive repertoire of highly standardized and/or formally validated assay platforms. This Core will also provide immune/molecular profiling leadership and expertise in collaboration with the Biostatistics and Bioinformatics Core (Core 1) and the Clinical Trials and Imaging Core (Core 2) for all projects. It is critical to the proposed projects to have well defined, accurately diagnosed high grade gliomas, a service that will be provided by this core. High quality blood and tissue samples needed for the proposed projects will be delivered by this core. Histopathologic services in this proposal include characterizing intratumoral heterogeneity in a spectrum of both well-differentiated and high grade infiltrating astrocytomas via tissue diagnosis and grading and in vitro analysis of antigenic expression in human glioma tissues as needed. The CMV pp65 antigen in GBM will be targeted and we will collaborate with the CLIA-certified Image Cytometry laboratory to develop a CMV CISH assay to characterize CMV presence and heterogeneity in high grade gliomas. Additionally, Core 3 will provide highly specialized and comprehensive assay platforms that include immunophenotyping, immune function analyses, and TCR sequencing. We will utilize high dimensional flow cytometry to determine if TReg depletion is sustained after anti-CD27 treatment in Project 1 and following lomustine administration in Project 3. We will use the polyfunctional T cell assay to determine if anti-CD27 increases the frequencies and relative polyfunctionality of CD4 and CD8 antigen-specific, vaccine-induced T cell responses. Assessment of T cell immune responses will also be measured using the same highly standardized and validated assays by this core in Project 2 and Project 3.